This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. We propose to investigate macro-ion induced pore formation in charged lipid membranes. Electrostatic interactions are known to be important for molecular "hole punchers" such as protein transduction domains that cross cell membranes, and membrane-active antimicrobial peptides that permeate cell membranes. The program concentrates on two cognate systems. (1) The HIV TAT protein transduction domain (PTD) is highly cationic arginine-rich peptide that can translocate across anionic eukaryotic cell membranes with anomalously high efficiency. We aim to explore arginine-rich, cell penetrating peptides, as well as synthetic analogs with well-defined sequences of arginines and other amino acids, in order to elucidate their mode of action on membranes.(2) Antimicrobial peptides (AMP) comprise a key component of innate immunity for a wide range of multicellular organisms. Theta-defensins are highly selective antimicrobial peptides. RTD-1 and BTD- 7 theta-defensins have activity against HIV and SIV. Protegrin (PG-1) is another antimicrobial peptide with 70% sequence homology to theta-defensins. However, protegrin is also active against some eukaryotic cells. We aim to understand how slight differences in amino acid composition and placement lead to large changes in peptide activity. Like protein transduction domains these cationic peptidesutilize the basic amino acid arginine over lysine and histidine. We investigate the self-assembled structure, interactions, and phase behavior of lipid vesicles in the presence of these two types of pore-forming agents using synchrotron x-ray diffraction. Success in this program of research can lead to advances across a broad range of biotechnological applications, such as improved drug delivery systems and design rules for antibiotics.